Field survival of containerized red and jack pine seedlings inoculated with mycelial slurries of ectomycorrhizal fungi

Red pine and jack pine seedlings growing in styroblocks were inoculated 8 wk after sowing with mycelium/agar slurries of 3 mycorrhizal fungi (Laccaria bicolor, Scleroderma citrinum, and an unidentified basidiomycete), and one suspected mycorrhizal fungus (Cantharellula umbonata). Seedlings inoculated with L. bicolor developed mycorrhizae earlier and in greater numbers than the other inoculation treatments, with red pine out-performing jack pine in both respects. At 34 wk following sowing, seedlings were outplanted on a cleared xeric site in Baraga Co., in Michigan's Upper Peninsula. Seedlings inoculated with C. umbonata failed to form mycorrhizae and were not outplanted. Inoculation treatments did not affect shoot or root weight at outplanting. Red pine inoculated with L. bicolor averaged 21% and 19% greater survival compared with control seedlings after one and two years in the field, respectively. Other inoculation treatments failed to increase seedling survival for either tree species. Jack pine demonstrated higher overall survival than did red pine for both years in all corresponding inoculation treatments.     

Ectomycorrhizal inoculation of Douglas-fir transplanted container seedlings with commercially produced inoculum

Commercially produced vegetative inocula of Laccaria laccata and Hebeloma crustuliniforme successfully formed ectomycorrhizae with Douglas-fir transplanted container (plug+1) seedlings. After 4.5 months in containers, 83% and 90%, respectively, of short roots were mycorrhizal. L. laccata- or H. crustuliniforme-inoculated seedlings had significantly more mycorrhizal and total short roots than Pisolithus tinctorius-inoculated (4% mycorrhizal root tips) or uninoculated control seedlings. No significant differences were detected in seedling growth at the end of the container phase.After transplantation and growth in nursery beds for 17 months, mean new short root colonization of all seedlings was 80%. H. crustuliniforme persisted as a dominant mycorrhizal fungus on seedlings initially inoculated with this fungus. L. laccata-inoculated seedlings had 40% of their short roots colonized by L. laccata and another 40% by native fungi Rhizopogon and Thelephora spp. All mycorrhizae of control seedlings and those inoculated with P. tinctorius were formed by fungi native to the nusery beds. A significant fungal treatment effect was detected for shoot height only. Control seedlings were significantly taller than L. laccata-inoculated seedlings after transplanting.This article is part of senior author's thesis in partial fulfillment of the Ph.D. degree in the Department of Forest Science at Oregon State University, Corvallis.     

蓝桉和尾叶桉混合菌根研究 Ⅱ.混合菌根的接种效应

接种苗木在生长量和生物量上均表现出极其显著的生长优势,ＥＣＭ真菌蜡蘑菌单接种及其与４种ＶＡ菌根菌剂混合接种,对苗木生长的促进作用尤为显著。在接种后１６周时,与对照苗相比,蓝桉和尾叶桉接种苗木的高生长量最大增幅分别为２８．８６％（ＬＳ）和８６．６５％（ＬＧ）;两种桉树地上部分平均干质量最大增幅分别为１２９．９３％（ＬＳ）和１３３．３４％（Ｌ）,地下部分分别为１１９．９３％（ＬＦ）和１７４．８３％（Ｌ     

Selection of efficient VA mycorrhizal fungi for Casuarina equisetifolia—second screening

An investigation was carried out to screen and select efficient vesicular arbuscular mycorrhizal (VAM) fungi for inoculating the forest tree species, Casuarina equisetifolia. The seedlings were inoculated with 10 different VAM fungi, obtained from various sources. Inoculated seedlings generally had greater plant height, stem girth, biomass and P content than uninoculated plants. They also had more mycorrhizal root colonization and spore numbers in root zone soil. C. equisetifolia seedlings responded best (in biomass) to inoculation with Glomus mosseae (Nicolson and Gerdemann) Gerdemann and Trappe, closely followed by Acaulospora laevis Gerdemann and Trappe and G. fasciculatum (Thaxter Sensu Gerdemann) Gerdemann and Trappe; all the three being statistically on par with each other.     

Effect of Scleroderma Spore Density and Age on Mycorrhiza Formation and Growth of Containerized Eucalyptus globulus and E. urophylla Seedlings

Greenhouse trials with containerized Eucalyptus globulus and E. urophylla seedlings were made to determine effective inoculum spore densities (Scleroderma cepa from temperate Australia, and S. citrinum from sub-tropical China) and spore storage conditions (fresh spores or spores stored for 5 years at room temperature or at 4 °C; S. albidum, S. areolatum and S. cepa from Western Australia). Inoculation with 10⁶ or 10⁸ spores/seedling increased eucalypt growth by up to 46% in height and 42% in dry weight compared to non-inoculated seedlings at 12 weeks after inoculation. Mycorrhizal formation was poor at 10² spores/seeding. Spores stored at 4 °C for 5 years were as effective in forming mycorrhizas as freshly collected spores when a standard density of 10⁶ spores/seedling was applied. It is recommended that spore densities ≥10⁴ be used for inoculation of containerised eucalypts and that spores be stored at 4 °C until use.     

Mycorrhizal dependency and growth responses of Acacia nilotica and Albizzia lebbeck to inoculation by indigenous AM fungi as influenced by available soil P levels in a semi-arid Alfisol wasteland

A series of available phosphorus (Olsen) levels ranging from 10 to 40 ppm were achieved in a semi-arid soil. The influence of the levels of phosphorus on the symbiotic interaction between two subtropical tree species, Acacia nilotica and Albizzia lebbeck, and a mixed inoculum of indigenous arbuscular mycorrhizal (AM) fungi was evaluated in a greenhouse study. The extent to which the plant species depended on AM fungi for dry matter production decreased as the levels of soil P increased, but the degree of this decrease differed in the two species tested. Acacia nilotica colonized by AM fungi showed a significant increase in shoot P and dry matter at a soil P level of 10 ppm whereas in Albizzia lebbeck, such increase occurred at 20 ppm. Mycorrhizal inoculation response disappeared beyond soil P levels of 25 ppm in Acacia nilotica and 30 ppm in Albizzia lebbeck. Levels of soil P greater than 25 ppm suppressed AM fungus colonization in both species. Soil P levels of 30 and 40 ppm and 40 ppm caused negative mycorrhizal dependencies (MD) in Acacia nilotica and Albizzia lebbeck respectively. Values of MD for both species were negatively correlated with soil P levels. Based on the MD values, regression equations were developed to predict MD for given levels of available P.     

Effect of post-planting inoculation with Bradyrhizobium sp and mycorrhizal fungi on the growth of Brazilian rosewood,Dalbergia nigra Allem. ex Benth., in two tropical soils

Six months after planting, seedlings of the leguminous treeDalbergia nigra, known as Brazilian rosewood, wereinoculated with a selected Bradyrhizobium strainBHICB-DN 15,either alone or in combination with mycorrhizal fungi to determine their effecton the growth of D. nigra in Atlantic forest andeucalyptussoils. Height growth of D. nigra was similar in both soilsand six months after inoculation Bradyrhizobium did notaffect the D. nigra height growth, but it did improve drymass and especially in the nitrogen content of plants grown in eucalyptus soil.The success of the delayed inoculation with BHICB-DN 15 confirms its competitiveability vis-à-vis indigenous soil rhizobia. Co-inoculationof BHICB-DN 15 and mycorrhizal fungi did not increase plant dry weight,nitrogenand phosphorus content or mycorrhizal colonization. These results suggest thatthe BHICB-DN 15 strain had no synergistic relationship with the mycorrhizalfungi or that there was incompatibility between symbionts, in both soils.     

Dual inoculation of a woody legume (Centrolobium tomentosum) with rhizobia and mycorrhizal fungi in south-eastern Brazil

The integration of N₂ fixing trees into stable agroforestry systems in the tropics is being tested due to their ability to produce high biomass N and P yields, when symbiotically associated with rhizobia and mycorrhizal fungi. The growth of Centrolobium tomentosum Guill. ex Benth, a native leguminous tree from the Brazilian Atlantic Forest, was assessed with dual inoculation of Rhizobium spp and mycorrhizal fungi under field conditions. Complete fertilization was compared to treatments of inoculation with selected rhizobia strains BHICB-Ab1 or BHICB-Ab3, associated or not to arbuscular mycorrhizal (AM) fungi. The dual inoculation increased the height and growth in relation to the plants treated with rhizobia alone. Plants inoculated with strain BHICB-Ab1 and arbuscular mycorrhizal fungi (AMF) exhibited an increase of 56% dry matter over uninoculated control and nitrogen accumulation was greater than with BHICB-Ab3 inoculated plants. Strain BHICB-Ab1 presented a synergetic relationship with mycorrhizal fungi since the combined inoculation with BHICB-Ab1 enhanced plant height and dry weight more than single inoculation while the growth of BHICB-Ab3 plants was not modified by AMF inoculation. Arbuscular mycorrhizal fungi enhanced plants survival and seemed to favour the nodule occupation by rhizobia strains as compared to the non-mycorrhizal plants. Inoculation with selected rhizobia and AMF improved the growth of C. tomentosum under field conditions. This revised version was published online in June 2006 with corrections to the Cover Date.     

Growth, nitrogen fixation and mineral acquisition of Alnus sieboldiana after inoculation of Frankia together with Gigaspora margarita and Pseudomonas putida

The role of tetrapartite associations among Frankia, Gigaspora margarita (an arbuscular mycorrhizal fungus), Pseudomonas putida (rhizobacterium), and Alnus sieboldiana in growth, nitrogen fixation, and mineral acquisition of A. sieboldiana was investigated. Seedlings of A. sieboldiana were inoculated with Frankia isolated from root nodules of alder, followed by inoculation of G. margarita and P. putida, and were grown for 5 months in a greenhouse. The seedlings inoculated with Frankia and G. margarita together produced the highest biomass of shoots and root nodules. Nitrogen-fixation activity, measured by acetylene reduction assay, was observed when Frankia was inoculated. The activity, on a per-nodule gram basis, decreased after G. margarita inoculation, but on a per-plant basis there was no significant difference in the activity among inoculation treatments. The mineral content in the seedlings changed after inoculation with Frankia, but not after inoculation with P. putida and/or G. margarita. The results showed a synergistic interaction among Frankia, the mycorrhizal fungus, and the rhizobacterium on the growth of A. sieboldiana.     

In vitro effects of four ectomycorrhizal fungi, Boletus edulis, Rhizopogon roseolus, Laccaria laccata and Lactarius deliciosus on Fusarium damping off in Pinus nigra seedlings

Species of Fusarium cause damping off of forest seedlings resulting in considerable losses and oftentimes fungicides are used to control the disease. Such chemicals may be detrimental to the environment. Protection of forest seedlings against this disease must be focused on integrated management for in which biological control is one of the most important tools. Mycorrhizal fungi may provide protection against pathogen invasion of seedlings. In this study, the interactions between the mycorrhizal fungi Boletus edulis, Rhizopogon roseolus, Laccaria laccata and Lactarius deliciosus and damping off pathogens (Fusarium oxysporum and F. moniliforme) were investigated in vitro and mechanisms involved in the protection of damping off in P. nigra seedlings are also discussed. The effect of filtrates from mycorrhizal species on spore germination of F. oxysporum varied throughout the tests. At the end of the assay, the inhibitory effect only could be observed in the L.␣deliciosus treatment. None of the filtrates reduced spore germination in F. moniliforme. Finally, three of the four mycorrhizal species grown for 2 months in the substrate, L. laccata, L.␣deliciosus and B. edulis, increased the survival of Fusarium inoculated P.␣nigra seedlings.     

Characterization and identification of mycorrhizas of Douglas fir in New Zealand

Thirteen types of natural Douglas fir (Pseudotsuga menziesii [Mirb.] Franco) mycorrhizas were characterized and nine of them were identified as P. menziesii + Rhizopogon sp., P. menziesii + R. vinicolor, P. menziesii + Hebeloma crustuliniforme, P. menziesii + Amanita muscaria, P. menziesii + Endogone flammicorona, P. menziesii + Boletus sp., P. menziesii + Laccaria laccata, P. menziesii + Thelephora terrestris and P. menziesii+Tuber sp.     

Effect of electric impulse on sporocarp formation of ectomycorrhizal fungusLaccaria laccata in Japanese red pine plantation

We examined the effect of electric impulses on sporocarp formations of ectomycorrhizal fungus in a Japanese red pine (Pinus densiflora) plantation.Laccaria laccata was the dominant species in this study plantation. Spatial distribution concentrated in the electric impulse treatment zone, and recorded a markedly larger number of sporocarps compared with untreated control plots. The effect of electric impulses was also recognized onL. laccata fruiting with a mycorrhizal sapling in a pot. Sporocarp production ofL. laccata was strongly promoted by the electric impulse in experiments bothin vivo natural plantation stands andin vitro pot saplings. Our work indicates the possibility of enhancement of sporocarp production of the mycorrhizal fungi with electric impulses in the nursery and plantation. This work was partly supported by a grant from the Kyushu Electric Power Company.     

Effect of Arbuscular Mycorrhiza on the Drought Tolerance of Poncirus trifoliata Seedlings

The effects of Glomus mosseae colonization on the plant growth and drought tolerance of 1-year-old trifoliate Poncirus trifoliata seedlings in potted culture were studied in natural water stress and rewatering conditions. Results showed that arbuscular mycorrhizal (AM) inoculation significantly improved the height, stem diameter, and fresh weight of P. trifoliata seedlings before natural water stress. By the end of the experiment, the survival percentage of AM-transplanted seedlings was 8% higher than those of non-AM ones. During water stress and rewatering, AM significantly increased the contents of soluble sugars and proteins in leaves, and enhanced the activities of superoxide dismutase (SOD), guaiacol peroxidase (G-POD), and catalase (CAT) in either seedling leaves or roots, which indicated that AM colonization could improve the osmotic adjustment response of P. trifoliata, enhance its defense system, and alleviate oxidative damages to membrane lipids and proteins. These results demonstrated that the drought tolerance of P. trifoliata seedlings was increased by inoculation with AM fungi. The functional mechanism underlying the observation that mycorrhizas increased the host’s drought tolerance was closely related to enzymatic and nonenzymatic antioxidant defense systems such as SOD, G-POD, CAT, and soluble protein. Translated from Chinese Journal of Applied Ecology, 2005, 16(3) (in Chinese)     

Effect of the inoculation and distribution of mycorrhizae in <Emphasis Type="Italic">Plathymenia reticulata</Emphasis> Benth under monoculture and mixed plantation in Brazil

This paper investigates the distribution of arbuscular mycorrhizal fungi (AMF) spores and AMF colonization in a field study in southeastern Brazil. Response to AMF and rhizobial inoculation was studied in monocultures of Plathymenia reticulata and mixed plantations with both Tabebuia heptaphylla and Eucalyptus camaldulensis in a sandy soil during two consecutive years. P. reticulata height and diameter and mycorrhizal colonization and AMF diversity were measured in dry and rainy periods. The inoculated treatment of E. camaldulensis, T. heptaphylla and P. reticulata mixed plants showed higher height and diameter growth of P. reticulata used as well as increased root colonization and AMF spore numbers. Spore populations were found to belong to five genera: Acaulospora, Entrophospora, Glomus, Gigaspora and Scutellospora, with Glomus dominating. Agroforestry practices including use of leguminous tree P. reticulata effectively maintained AMF spore numbers in soils and high AMF colonization levels compared with monocultures, proving an efficient system for productivity and sustainability.     

Soil factors influencing the growth response of <Emphasis Type="Italic">Acacia holosericea</Emphasis> A. Cunn. ex G. Don to ectomycorrhizal inoculation

Australian acacias like Acacia holosericea are excellent candidates for the revegetation of arid zones in Africa. Their high ability to develop multiple symbioses with soil microorganisms is crucial to their rapid development in adverse climatic and edaphic conditions. These symbioses include nitrogen fixation with rhizobia, vesicular arbuscular mycorrhization and ectomycorrhization. We set up an experiment to test the growth response of A. holosericea to ectomycorrhizal inoculation in 13 different soils from Senegal. After autoclaving the soil, the experiment was conducted in a greenhouse for four weeks. Plants were inoculated with Pisolithus albus strain IR100. The following parameters were measured: plant biomass, N, P, K and Ca foliar composition, spontaneous nodulation rate, and ectomycorrhizal colonization. Data were analyzed in light of the physical, chemical and total microbial characteristics of the soil. The results demonstrated a global promoting effect of P. albus inoculation on plant (shoot and root) growth and on foliar P and K, together with a depressive effect on N, while Ca rates were barely affected. Interestingly, spontaneous nodulation with putative water- or airborne rhizobia was stimulated after P. albus inoculation. However, these nodules seemed poorly effective, as they failed to cause any change in plant growth or in foliar N composition within the nodulated and nonnodulated control plants. These results show that plant growth response to ectomycorrhizal inoculation is greatly dependent on soil characteristics, and that root ectomycorrhizal colonization is influenced by biotic factors such as soil microbiota. From a practical point of view, data from the present study demonstrate that it is possible to optimize the effect of fungi on plant growth by screening soils under nursery conditions.     

Vesicular-arbuscular mycorrhiza effects on Gliricidia sepium and Senna siamea in a fallowed alley cropping system

Nitrogen fixing and non-N₂ fixing legumes such as Gliricidia speium and Senna siamea have been used in alley cropping systems for soil improvement and source of N for food crops. However their establishments could be limited by P and moisture deficiencies in degraded soils. Vesicular-arbuscular mycorrhizal fungi can help to overcome these deficiencies. We examined the effects of a vesicular-arbuscular mycorrhizal (VAM) fungus, Glomus deserticola, on the performance of sole hedgerow trees of Gliricidia sepium and Senna siamea and their mixtures (interplanted) in a fallowed alley cropping experiment on a degraded Alfisol in southwestern Nigeria. Percentage root infection by VAM fungi was higher in inoculated plants than in uninoculated ones irrespective of whether they were interplanted or non-interplanted. Inoculation with G. deserticola increased dry matter accumulation and nutrient uptake (N. P, Mg and K) but there was no significant interaction between mycorrhizal inoculation and interplanting for growth and nutrient uptake except for the uptake of P, Mg and K in G. sepium. Inoculation with G. deserticola reduced leaf shedding of G. sepium by 50% but did not have the same effect for S. siamea. For both tree species inoculated plants extracted more water from 0–30 cm depth than the uninoculated ones.     

蓝桉和尾叶桉混合菌根研究Ⅱ.混合菌根的接种效应

接种苗木在生长量和生物量上均表现出极其显著的生长优势,ECM真菌蜡蘑菌单接种及其与4种VA菌根菌剂混合接种,对苗木生长的促进作用尤为显著。在接种后16周时,与对照苗相比,蓝桉和尾叶桉接种苗木的高生长量最大增幅分别为28.86%(LS)和86.65%(LG);两种桉树地上部分平均干质量最大增幅分别为129.93%(LS)和133.34%(L),地下部分分别为119.93%(LF)和174.83%(LG)。桉树对不同菌根菌及其组合的菌根依赖性表现出很大的差异,对混合接种的真菌组合有相对高的菌根依赖性,而对土壤菌剂依赖性很小。菌根感染与苗木生物量间有显著的相关性。试验结果还证实了P素对桉苗生长的促进作用,P素水平对菌根的接种效应有一定的影响。     

Effects of single and dual inoculation with selected microsymbionts (rhizobia and arbuscular mycorrhizal fungi) on field growth and nitrogen fixation of <Emphasis Type="Italic">Calliandra calothyrsus</Emphasis> Meissn

Our study aimed to assess the effect of dual inoculation with the selected Rhizobium strain KWN35 and the arbuscular mycorrhizal isolate BEG 176 (Glomus etunicatum) on the growth of Calliandra calothyrsus cultivated under irrigation in the field in Senegal (Dakar) over a period of 24 months. Although plants inoculated with both microsymbionts grew better (height and root collar diameter) than plants from three other inoculation treatments (control, single inoculation with KWN35 or BEG 176), these results were not statistically different except at 1 and 5 months after field transplantation. KWN35 was present in a relatively high percentage of nodules harvested from plants inoculated with this rhizobial strain either alone or with mycorrhiza (approximately 60% and 40% 1 and 2 years after field transplantation respectively). The percentage of mycorrhizal root infections was around 60% in the inoculated plants after 12 months with significantly higher N, P and K foliar contents of trees compared with the non-inoculated controls. However, as for nodulation, by 24 months after transplantation, there were no significant differences between treatments. We conclude that field inoculation of C. calothyrsus with Rhizobium strain KWN35 and arbuscular mycorrhizal isolate BEG 176 did not have a long-lasting effect on the growth of trees, even when a majority of nodules were occupied by the inoculated rhizobia and the roots infected by the mycorrhiza. Several reasons can be postulated for these results, such as a possible effect of soil fertility on the efficiency of the nodules.     

Seed source and vesicular-arbuscular mycorrhizal symbiont affects growth of Juglans nigra seedlings

Seedlings of Juglans nigra from three seed sources were grown in fumigated soil without vesicular-arbuscular mycorrhizal (VAM) fungi or inoculated with Gigaspora margarita, Glomus deserticola or Glomus etunicatum. Vesicular-arbuscular mycorrhizal development varied significantly between fungal symbionts within a black walnut source. Glomus deserticola and G. etunicatum produced the highest levels of root colonization in all sources. Significant differences in seedling shoot and root growth were attributed to root colonization by specific VAM fungi in each black walnut source. Glomus deserticola stimulated seedling leaf area and root weight 26 and 52%, respectively, in one seed source. Seedling leaf N, P and K concentrations were significantly improved by VAM in two seed sources. Juglans nigra seedlings respond favorably to VAM colonization. However, differences between seed sources suggest a strong host-symbiont interaction.     

Procedure for establishing sheathing mycorrhizas on tree seedlings

A simple procedure is described where seedlings emerging from surface sterilised seeds (of Betula pendula, B. pubescens, Pinus contorta and Picea sitchensis) were grown axenically in a peat/vermiculite mixture which had been previously inoculated, and incubated, with sheating mycorrhizal fungi (including Hebeloma sacchariolens, Paxillus involutus and Amanita muscaria). Mostly, seedlings were grown in split polystyrene tubes (50 mm tall x 13 mm internal diameter) which were arranged $\text{140}\text{150}$ per tray, the trays being covered with transparent propagator lids so forming a unit that could be sealed and sterilised before use with gamma radiation. With the formation of sheating mycorrhizas after about 7 weeks the sterility of the container was broken to enable seedlings to be hardened off prior to transplanting.Roots of Betula pendula seedlings grown in tubes were smaller, as judged by numbers of ‘fragments’, than those of plants which were not ‘restricted’. ‘Tubing’ increased: (a) the proportion of root fragments with mycorrhizas; and (b) the sizes of individual mycorrhizas. The sizes of the ‘tubing’ effects differed with different isolates of A. muscaria, so suggesting, because of the sensitivity of the system, that environmental and genotypic factors must be carefully controlled.